The major objective of this research proposal is to define the mechanisms of insulin resistance in various altered physiologic states in man and to elucidate the relationships between measurements of in vivo and in vitro insulin action. We propose to apply this combined in vivo and in vitro approach to the insulin resistance of human obesity, non-ketotic diabetes mellitus, and aging. Additionally, we will assess the effects of clinically relevant perturbations such as high carbohydrate feedings and treatment with oral sulfonylurea agents. With this approach we will determine under what conditions insulin resistance is directly related to decreased insulin receptors and under what conditions post-receptor defects are implicated. In vivo studies will be carried out by use of the glucose clamp technique. In vitro studies will involve measurements of insulin binding to receptors on isolated adipocytes, monocytes and erythrocytes. In addition, in vitro measurements of glucose transport in freshly isolated adipocytes will be performed, and studies of insulin binding and glucose transport in cultured fibroblasts will also be carried out. The in vitro studies of insulin function and glucose metabolism will allow a quantitative correlation of in vivo and in vitro data and will provide direct answers as to which specific cellular defects exist. Lastly, the use of a freshly isolated in vitro system (adipocytes) and a cultured cell system (fibroblasts) should allow us to distinguish between primary cellular defects as opposed to defects which are secondary to some aspect of the in vivo environment. It is expected that the studies proposed will provide information and test hypotheses which should help elucidate the causes of insulin resistance in various human disease states. Clearly, a better understanding of the mechanisms of insulin resistance can potentially lead to a more rational design and understanding of therapeutic approaches.